Method of treating high plasma cholesterol levels

ABSTRACT

A method prevents or treats diseases associated with high plasma cholesterol levels. In addition, this method reduces plasma cholesterol levels. The method comprises administering a lipase inhibitor, e.g. orlistat, and a pharmaceutically acceptable bile acid sequestrant.

[0001] This application is a divisional of U.S. patent application Ser.No. 09/912,956, filed Jul. 25, 2001, currently pending.

BACKGROUND OF THE INVENTION

[0002] 1. Field

[0003] The present invention relates to a new method for the preventionand treatment of diseases associated with high plasma cholesterol levels(hypercholesterolemia).

[0004] 2. Description

[0005] Bile acid sequestrants have been proposed for use in loweringcirculating blood cholesterol levels. Biologically, cholesterol iseliminated from the body by conversion to bile acids and excretion asneutral steroids. Bile acids are synthesized from cholesterol in theliver and enter the bile as glycine and taurine conjugates. They arereleased in salt form in bile during digestion and act as detergents tosolubilize and consequently aid in digestion of dietary fats. Followingdigestion, bile acid salts are mostly reabsorbed in the ileum, complexedwith proteins, and returned to the liver through hepatic portal veins.The small amount of bile acid salts which are not reabsorbed by activetransport are excreted via the distal ileum and large intestine as aportion of fecal material. Bile acids are synthesized from cholesteroltransported in lipoproteins in the liver. Therefore, reabsorption ofbile acids, which can be present as the corresponding salts orconjugates, from the intestine conserves lipoprotein cholesterol in thebloodstream. As such, reducing reabsorption of bile acids within theintestinal tract can lower levels of bile acid circulating in theenterohepatic system thereby promoting replacement of bile acids throughsynthesis from cholesterol, in the liver. The result is a lowering ofcirculating blood cholesterol levels. One method of reducing the amountof bile acids that are reabsorbed, is oral administration of compoundsthat sequester the bile acids within the intestinal tract and cannotthemselves be absorbed. The sequestered bile acids consequently areexcreted.

[0006] Orlistat (also known as tetrahydrolipstatin and sold under thetrademark XENICAL®) is a potent inhibitor of gastrointestinal lipases,i.e. lipases that break down ingested fat (gastric lipase, carboxylesterlipase, pancreatic lipase). Orilistat has the formula:

[0007] As a consequence of gastrointestinal lipase inhibition,unabsorbed fat is egested in the faeces. Pancreatic lipase is the keyenzyme for the hydrolysis of dietary triglycerides. Triglycerides, whichhave escaped hydrolysis are not absorbed in the intestine. Orlistat hasbeen shown in animal models to inhibit fat absorption and to reduce bodyweight. In pharmacological studies with humans potent inhibition of fatabsorption was demonstrated.

[0008] Hypercholesterolemia is a great problem in modern society.Accordingly, methods for treating hypercholesterolemia are highlysought.

SUMMARY OF THE INVENTION

[0009] The subject invention provides a method of reducing plasmacholesterol levels in a hypercholesterolemic patient. This methodcomprises administering to the patient a therapeutically effectiveamount of a lipase inhibitor and a therapeutically effective amount of abile acid sequestrant. A favored lipase inhibitor is orlistat andpreferred bile acid sequestrants include cholestyramine, colestipol,colesevelam, colestimide, sevelamer, cellulose derivatives, dextranderivatives, starch, starch derivatives, and pharmaceutically acceptablesalts of such bile acid sequestrants. A key advantage of the subjectinvention is the mitigations of the side effects associated with bothorlistat (oily stool) and bile acid sequestrants (constipation).

[0010] The bile acid sequestrant can be a cellulose derivative or adextran derivative, for example DEAE-cellulose, guanidinoethylcellulose,or DEAE-Sephadex. Favored starch derivatives include β-cyclodextrin,γ-cyclodextrin, retrograded starch, degraded starch, a mixture ofretrograded and degraded starch, hydrophobic starch, amylose,starch-diethylaminoethylether, and starch-2-hydroxyethylether.β-cyclodextrin or γ-cyclodextrin are preferred.

[0011] The bile acid sequestrant is favorably cholestyramine,colestipol, colesevelam, colestimide, sevelamer, cellulose,DEAE-cellulose, guanidinoethylcellulose, DEAE-Sephadex, starch,β-cyclodextrin, and γ-cyclodextrin. More favored are cholestyramine,colestipol, colesevelam, colestimide, sevelamer, DEAE-cellulose,β-cyclodextrin, and γ-cyclodextrin, and in particular, cholestyramine,colestipol, sevelamer, DEAE-cellulose, β-cyclodextrin, andγ-cyclodextrin. Presently it is preferred that the bile acid sequestrantis cholestyramine or colestipol or sevelamer.

[0012] It is favored when the therapeutically effective amount of alipase inhibitor and a therapeutically effective amount of a bile acidsequestrant are administered as a medicament that comprises about 5 mgto about 1000 mg lipase inhibitor, about 0.1 g to about 20 g bile acidsequestrant, and one or more pharmaceutically acceptable excipients. Thepharmaceutically acceptable excipients are typically fillers, sugars,sugar alcohols, a mixture of one or more sugars and sugar alcohols,surfactants, disintegrants, binders, lubricants, flowability enhancers,sweeteners, and colorants.

[0013] Another favored variation is where the medicament comprises:about 5 mg to about 1000 mg lipase inhibitor; about 0.1 g to about 20 gbile acid sequestrant; and a pharmaceutically acceptable excipientsselected from the group consisting of about 0.1 to about 10 g fillers,sugars, sugar alcohols and mixtures thereof, about 0.05 g to about 3.0 gsurfactants, about 0.05 g to about 2.0 g disintegrants, about 0.02 g toabout 2.0 g binder, about 0.001 g to about 1.0 g lubricants, about 0.1 gto about 5.0 g flowability enhancers, about 0.01 g to about 4.0 gsweeteners, and about 0.001 g to about 0.5 g colorants. Such medicamentstypically contain about 10 mg to about 500 mg lipase inhibitor, with therange about 20 mg to about 100 mg lipase inhibitor being favored.Favored is where the medicament comprises about 10 mg to about 360 mgorlistat, 30 mg to about 120 mg orlistat being favored and about 40 mgto about 80 mg orlistat being most favored.

[0014] The medicament generally comprises about 0.5 g to about 10 g bileacid sequestrant, although about 1 g to about 5 g bile acid sequestrantis favored. Especially favored is where the bile salt sequestrant ischolestyramine.

[0015] Administration of the lipase inhibitor and bile acid sequestrantcan be simultaneous, sequential, and/or separate.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0016] The subject invention will now be described in terms of itspreferred embodiments. These embodiments are set forth to aid inunderstanding the invention but are not to be construed as limiting.

[0017] Obesity is often associated with cardiovascular risk factors,especially with elevated levels of plasma cholesterol. A clinicallyestablished principle to treat elevated cholesterol levels is theadministration of bile acid sequestrants. One side effect of bile acidsequestrants is constipation. The present invention provides improvedmethods for the prevention and treatment of diseases associated withoverweight and high plasma cholesterol levels.

[0018] The present invention provides the combined use of a lipaseinhibitor and a pharmaceutically acceptable bile acid sequestrant forthe manufacture of medicaments for the prevention and treatment ofdiseases associated with high plasma cholesterol levels. Surprisingly,the combination results in both improved cholesterol lowering andattenuation of the side effects seen with either administration oflipase inhibitors or bile acid sequestrants individually.

[0019] In more detail, the present invention relates to the use of alipase inhibitor, e.g. orlistat, and a pharmaceutically acceptable bileacid sequestrant for the manufacture of a medicament for the preventionand treatment of diseases associated with high plasma cholesterollevels, i.e. for the treatment of hypercholesterolemia. Preferably, thepharmaceutically acceptable bile acid sequestrant is selected from thegroup consisting of cholestyramine, colestipol, colesevelam,colestimide, sevelamer, cellulose and dextran derivatives, starch andstarch derivatives and pharmaceutically acceptable salts thereof.

[0020] Unless otherwise indicated the following definitions are setforth to illustrate and define the meaning and scope of the variousterms used to describe the invention herein.

[0021] The term “pharmaceutically acceptable” as used herein means thatthe buffer or salts are acceptable from a toxicity viewpoint.

[0022] The term “pharmaceutically acceptable salts” as used herein meanssalts of lipase inhibitors or bile acid sequestrants with inorganic ororganic acids such as hydrochloric acid, hydrobromic acid, sulphuricacid, phosphoric acid, citric acid, formic acid, maleic acid, aceticacid, succinic acid, tartaric acid, methanesulphonic acid,p-toluenesulphonic acid and the like, which are non-toxic to livingorganisms. In the case of bile acid sequestrants having quaternaryammonium groups pharmaceutically acceptable salts mean correspondinglychlorides, bromides, sulphates, phosphates, citrates, formiates,maleates, acetates, succinates, tartrates, methanesulphonates,p-toluenesulphonates and the like.

[0023] The term “lipase inhibitor” refers to compounds which are capableof inhibiting the action of lipases, for example gastric and pancreaticlipases. For example orlistat and lipstatin as described in U.S. Pat.No. 4,598,089 are potent inhibitor of lipases. Lipstatin is a naturalproduct of microbial origin, and orlistat is the result of ahydrogenation of lipstatin. Other lipase inhibitors include a class ofcompound commonly referred to as panclicins. Panclicins are analogues oforlistat (Mutoh et al, 1994). The term “lipase inhibitor” refers also topolymer bound lipase inhibitors for example described in InternationalPatent Application WO99/34786 (Geltex Pharmaceuticals Inc.). Thesepolymers are characterized in that they have been substituted with oneor more groups that inhibit lipases. The term “lipase inhibitor” alsocomprises pharmaceutically acceptable salts of these compounds. The term“lipase inhibitor” also refers to 2-oxy-4H-3,1-benzoxazin-4-ones whichhave been described in International Patent Application WO00/40569(Alizyme Therapeutics Ltd.), e.g.2-decyloxy-6-methyl-4H-3,1-benzoxazin-4-one,6-methyl-2-tetradecyloxy-4H-3,1-benzoxazin-4-one, and2-hexadecyloxy-6-methyl-4H-3,1-benzoxazin-4-one. Most preferably, theterm “lipase inhibitor” refers to orlistat.

[0024] Orlistat is a known compound useful for the control or preventionof obesity and hyperlipidemia. See, U.S. Pat. No. 4,598,089, issued Jul.1, 1986, which also discloses processes for making orlistat and U.S.Pat. No. 6,004,996, which discloses appropriate pharmaceuticalcompositions. Further suitable pharmaceutical compositions are describedfor example in International Patent Applications WO 00/09122 and WO00/09123. Additional processes for the preparation of orlistat aredisclosed in European Patent Applications Publication Nos. 185,359,189,577, 443,449, and 524,495.

[0025] Orlistat is preferably orally administered from 60 to 720 mg perday in divided doses two to three times per day. Preferred is whereinfrom 180 to 360 mg, most preferably 360 mg per day of a lipase inhibitoris administered to a subject, preferably in divided doses two or,particularly, three times per day. The subject is preferably an obese oroverweight human, i.e. a human with a body mass index of 25 or greater.Generally, it is preferred that the lipase inhibitor be administeredwithin about one or two hours of ingestion of a meal containing fat.Generally, for administering a lipase inhibitor as defined above it ispreferred that treatment be administered to a human who has a strongfamily history of obesity and has obtained a body mass index of 25 orgreater.

[0026] Orlistat can be administered to humans in conventional oralcompositions, such as tablets, coated tablets, hard and soft gelatincapsules, emulsions or suspensions. Examples of carriers which can beused for tablets, coated tablets, dragees and hard gelatin capsules arelactose, other sugars and sugar alcohols like sorbitol, mannitol,maltodextrin, or other fillers; surfactants like sodium lauryl sulfate,Brij 96, or Tween 80; disintegrants like sodium starch glycolate, maizestarch or derivatives thereof; polymers like povidone, crospovidone;talc; stearic acid or its salts and the like. Suitable carriers for softgelatin capsules are, for example, vegetable oils, waxes, fats,semi-solid and liquid polyols and the like. Moreover, the pharmaceuticalpreparations can contain preserving agents, solubilizers, stabilizingagents, wetting agents, emulsifying agents, sweetening agents, coloringagents, flavoring agents, salts for varying the osmotic pressure,buffers, coating agents and antioxidants. They can also contain stillother therapeutically valuable substances. The formulations mayconveniently be presented in unit dosage form and may be prepared by anymethods known in the pharmaceutical art. Preferably, orlistat isadministered according to the formulation shown in the Examples and inU.S. Pat. No. 6,004,996, respectively.

[0027] The term “bile acid sequestrant” refers to compound classes whichare able to bind bile acids or bile acid salts by various principles,e.g. anion exchanging polymers containing amino groups, quaternaryammonium groups, etc. (amine containing polymers). In more detail theterm refers to oligomers or polymers of different architecture (graft-,block-, multiblock-, homo-, copolymers), dendrimers, or hyperbranchedstructures containing either quaternary ammonium groups, substituted orunsubstituted pyridinium groups, substituted or unsubstituted primary,secondary, or tertiary alkyl- or arylamine groups, or any statistical ornon-statistical combination thereof, which are capable of formingcomplexes with physiologically active bile acids and/or bile saltsthrough non-covalent Van der Waals, hydrophobic and/or ionicinteractions. For example, these structures can include, e.g.,poly(amino acids) such as poly(lysine), poly(lactic acid-co-lysine)(PLAL), poly(vinyl amine), poly(allyl amine), poly(N-alkylvinyl amine),poly(N,N-dialkyl amine), poly(N-alkylallyl amine), poly(ethylene imine)and other mono- or disubstituted poly(amine)s. Further polymers includepoly(vinyl pyridinyl), poly(amide enamines), PAMAM dendrimers, polymerscontaining azo-groups, poly(dialkyl siloxane)s, poly(phosphazene)s,poly(acrylate)s, poly(methacrylate)s, poly(styrene), poly(amides),poly(ethers), poly(esters). Suitable side-chains can include cationic orneutral groups, substituted and unsubstituted alkyl or aryl groups,saturated or unsaturated alkyl groups, amino acids or functional groupssuch as amine or ammonium moieties, for example (Uhrich et. al., Chem.Rev. 1999, 99, 3181-3198). In addition, naturally occurring andsubsequently synthetically modified polymers such as poly(aminosaccharide)s (chitosan) or cellulose derivatives (e.g.diethylaminoethylcellulose, guanidinoethylcellulose) are also ofparticular interest. A further important class of bile acid sequestrantsare compounds capable of forming host-guest inclusion complexes, such asβ and γ-cyclodextrines.

[0028] Bile acid sequestrants and methods for their preparation havebeen described for example in International Patent ApplicationsWO95/34585 (Geltex Pharmaceuticals, Inc.; relating to polyamine salthydrophobic sequestrants), WO94/27620 (Geltex Pharmaceuticals, Inc.;relating to the preparation of polymeric sequestrants for bile acids),and WO94/04596 (DuPont; relating to crosslinked polymeric ammoniumsalts).

[0029] For example, amine containing polymers, as defined herein, maycomprise the compounds described in International Patent ApplicationWO94/27620. The polymers are characterized by a repeat unit having theformula

[0030] or copolymer thereof, where n is an integer; R¹ is H or an alkylgroup (which may be straight chain or branched, substituted orunsubstituted, e.g., a C₁-C₈ alkyl, such as methyl); M is —C(O)—R² or-Z-R²; Z is O, NR³, S, or (CH₂)_(m); m=0-10; R³ is H or an alkyl group(which may be straight chain or branched, substituted or unsubstituted,e.g., C₁-C₈ alkyl, such as methyl); and R² is

[0031] where p=0-10, and each R⁴, R⁵, and R⁶, independently, is H, analkyl group (which may be straight chain or branched, substituted orunsubstituted, e.g., C₁-C₈ alkyl, such as methyl), or an aryl group(e.g., having one or more rings and which may be substituted orunsubstituted, e.g., phenyl, naphthyl, imidazolyl, or pyridyl). Inpreferred embodiments, the polymer is crosslinked by means of amultifunctional crosslinking co-monomer, the co-monomer being present inan amount from about 0.5-25% (more preferably about 2.5-20% (or about1-10%)) by weight, based upon total monomer weight. The compositions arenon-toxic and stable when ingested in therapeutically effective amounts.The preparation of these compounds is described in International PatentApplication WO94/27620 (Geltex Pharmaceuticals Inc.)

[0032] The term “bile acid sequestrants” also refer to compoundsobtainable by molecular imprinting. Molecular imprinting is based ontemplate polymerization in which polymers are prepared in the presenceof a template molecule to be imprinted. Resulting polymers record theshapes and chemical properties of the templates in their matrixes andexhibit specific binding characteristics to the template molecule. Themost significant advantage of this technique is to provide a simpleprocedure for the preparation of synthetic polymers capable of molecularrecognition: complementary sites for the target molecules which areexpected to be specific binding sites can be constructed with the aid ofmolecular self-assembling and no complicated synthesis is necessary.Molecular imprinted polymers have been described and the effectivenessof molecular imprinted polymers have been demonstrated (Ansell et al.Curr. Opin. Biotechnol. (1996), 7(1), 89-94). Imprinted polymers havebeen used for the chromatographic separation of amino acids, sugars,drugs and nucleotides. Drugs have been measured using imprinted polymersas antibody substitutes in radioligand binding assays (Sheaet al. TrendsPolym. Sci. (1994), 2(5), 166-73; Takeuchi et al. Chromatography (1997),18(2), 102-103; Nicholls, J. Molecular Recognition, (1988) 11 (1-6),79-82). The term “bile acid sequestrants” also comprisespharmaceutically acceptable salts of these compounds.

[0033] More preferably the invention refers to compositions orcombinations wherein the pharmaceutically acceptable bile acidsequestrant is selected from the group consisting of cholestyramine,colestipol, colesevelam, colestimide, sevelamer, cellulose and dextranderivatives, starch and starch derivatives and pharmaceuticallyacceptable salts thereof.

[0034] Cholestyramine (Quantalan®, Bristol-Myers Squibb) is a knowncompound and described for example in U.S. Pat. No. 4,902,501 and thereferences cited therein. It is a strong cationic resin containingquaternary ammonium functional groups bonded to a polymericstyrene-divinylbenzene structure:

[0035] It was introduced in the therapy in 1959 and is prevailingly usedin the management of the hypercholesterolemic states. The therapeuticalactivity of colestyramine is correlated to the capability of that resinof sequestering the biliary acids of the intestine, increasing up to 7-8times the faecal elimination thereof. The use of cholestyramine resin asadjunctive therapy to diet in the management of patients with elevatedcholesterol levels is noted in Remington's Pharmaceutical Sciences, 15thEd. Mack Publishing Co. (1975) pp 733-734. Methods for the preparationof cholestyramine and appropriate compositions are known in the art(e.g. DE-A-38 08 191, EP-A-347 014, U.S. Pat. Nos. 5,695,749, 4,172,120and EP-A-492 235).

[0036] Colestipol (Cholestabyl®, Pharmacia & Upjohn) is a known compoundand described for example in U.S. Pat. Nos. 3,692,895, 3,803,237, and5,807,582 and the references cited therein. It is a basic anion exchangeresin described as a high molecular weight copolymer ofdiethylenetriamine and 1-chloro-2,3-epoxypropane (epichlorohydrin), withapproximately one out of 5 amine nitrogens protonated, i.e. it is acopolymer of diethylenetriamine and epichlorohydrin with about 1 out of5 amine nitrogens protonated:

[0037] It is a light yellow resin which is hygroscopic and swells whenplaced in water or aqueous fluids. See Merck Index (Tenth Edition)#2440, page 2438. Colestipol hydrochloride is commercially available ingranule form as Colestid® Granules. See Physicians Desk Reference (PDR)42nd Ed., p. 2119 (1988). Colestid® Granules are marketed as ahyperlipidemia agent for oral use. Colestipol binds bile acids in theintestine forming a complex that is excreted in the faeces. Thisnonsystemic action results in a partial removal of the bile acids fromthe enterohepatic circulation, preventing their reabsorption.

[0038] Colesevelam and colesevelam hydrochloride (Cholestagel® orWelChol®) are cholesterol-lowering agents (Polym. Prepr. 2000, 41,735-736). Colesevelam is a polyamine-copolymer of the three aminesmentioned below, which are cross-linked with epichlorhydrine. Othernames are 1-Hexanaminium, N,N,N-trimethyl-6-(2-propenylamino)-,chloride, polymer with (chloromethyl)oxirane, 2-propen-1-amine andN-2-propenyl-1-decanamine, hydrochloride (9CI); or 1-Decanamine,N-2-propenyl-, polymer with (chloromethyl)oxirane, 2-propen-1-amine andN,N,N-trimethyl-6-(2-propenylamino)-1-hexanaminium chloride,hydrochloride (9CI); 2-Propen-1-amine, polymer with(chloromethyl)oxirane, N-2-propenyl-1-decanamine andN,N,N-trimethyl-6-(2-propenylamino)-1-hexanaminium chloride,hydrochloride (9CI); Oxirane, (chloromethyl)-, polymer with2-propen-1-amine, N-2-propenyl-1-decanamine andN,N,N-trimethyl-6-(2-propenylamino)-1-hexanaminium chloride,hydrochloride (9CI); Cholestagel; Colesevelam hydrochloride; GT 31-104;or GT 31-104HB (see also Holmes-Farley, S. et al.; Polym. Prepr. (Am.Chem. Soc., Div. Polym. Chem.) (2000), 41(1), 735-736. The threerelevant amines have the formula

[0039] Epichlorohydrine has the following formula

[0040] Colesevelam and colesevelam hydrochloride have been described inU.S. Pat. Nos. 5,607,669; 5,624,963; 5,679,717; 5,693,675; 5,917,007;and 5,919,832:

[0041] Sevelamer and its hydrochloride (Renagel®, GelTex) is a polymericphosphate binder intended for oral administration. Sevelamerhydrochloride is poly(allylamine hydrochloride) crosslinked withepichlorohydrin in which forty percent of the amines are protonated:

[0042] It is known chemically aspoly(allylamine-co-N,N′-diallyl-1,3-diamino-2-hydroxypropane)hydrochloride. Sevelamer hydrochloride is hydrophilic, but insoluble inwater. The compound, its preparation and use has been described in U.S.Pat. Nos. 5,496,545 and 5,667,775 and in International PatentApplication WO95/05184.

[0043] Colestimide (Cholebine®; Mitsubishi-Tokoyo Pharmaceuticals) is a2-methylimidazole polymer with 1-chloro-2,3-epoxypropane:

[0044] The binding of bile acids has been described e.g. in Mitsuka etal. Japanese Pharmacology & Therapeutics, 24 (Suppl. 4), 103, 1996,Mitsuka et al. Japanese Pharmacology & Therapeutics, 24 (Suppl. 4), 111,1996 and Mitsuka et al. Japanese Pharmacology & Therapeutics, 24 (Suppl.4), 127, 1996.

[0045] Various cellulose and dextran anion exchangers bind bile acids invitro under conditions of pH and ionic strength resembling those in thelumen of the small intestine (Parkinson, Thomas M., J. Lipid Res.(1967), 8(1), 24-9; Nichifor et la. Pharma Sci. (1994), 4(6), 447-52).Of these substances, DEAE-cellulose, guanidinoethylcellulose, andDEAE-Sephadex reduce hypercholesterolemia when added to the diet ofcholesterol-fed cockerels. In addition, DEAE-Sephadex reduced serumsterols in normocholesterolemic cockerels and dogs, lowered serumphospholipids and triglycerides in cholesterol-fed hypercholesterolemiccockerels and in normocholesterolemic dogs, and increased fecalexcretion of bile acids in hypercholesterolemic cockerels. Theseinsoluble cationic polymers evidently exert their hypocholesterolemiceffects by interrupting the enterohepatic circulation of bile acids.DEAE-celulloses are compounds wherein diethylaminoethyl-groups arecovalently bound to the cellulose hydroxylic groups. DEAE-celluloses areknown substances and commercially available (e.g. Sigma-Aldrich).Guanidinoethylcelluloses are compounds wherein guanidinoethyl-groups arecovalently bound to the cellulose hydroxylic groups.Guanidinoethylcelluloses are known substances and commerciallyavailable. DEAE-Sephadex is a cross-linked dextran derivative whereindiethylaminoethyl-groups are covalently bound to dextran. DEAE-Sephadexcompositions are commercially available (e.g. Pharmacia FinceChemicals). DEAE-cellulose, guanidinoethylcellulose and DEAE-Sephadexare especially useful as bile sequestrant agent, preferablyDEAE-cellulose.

[0046] The term “starch and derivatives thereof” comprise compoundswhich are able to form inclusion complexes with free and conjugated bilesalts and bile acids. Examples are β-, and γ-cyclodextrin, whichcontain, respectively, seven, and eight anhydroglucose (C₆H₁₀O₅) units.These molecules are doughnut-shaped rings having a hollow cavity of aspecific volume. The polar hydroxyl groups are oriented to the outsideof the rings, giving the outer surface a hydrophilic nature. Incontrast, the internal cavity has a hydrophobic (lipophilic) nature.Because of this unique structure, cyclodextrins, as the “host”molecules, are able to hold “guest” molecules of suitable size(generally of a molecular weight between 80 and 250), shape, andhydrophobicity within their cavity. (“Production and Potential FoodApplications of Cyclodextrins” Food Technology, January 1988, pp.96-100). β-, and γ-cyclodextrin are commercially available compounds(e.g. Sigma-Aldrich). Other examples of starch and derivatives thereofare retrograded and/or degraded starch e.g. maltodextrin, hydrophobicstarch, amylose, and starch derivatives, e.g.starch-diethylaminoethylether, starch-2-hydroxyethylether and the like(“Lexikon der Hilfsstoffe für Pharmazie, Kosmetik und angrenzendeGebiete”, H. P. Fiedler, Editio Cantor Aulendorf, Bd. 2., 3. Auflage,1989, pp. 1147-1154). Preferably starch derivatives are selected fromthe group consisting of consisting of β- or γ-cyclodextrin, retrogradedand/or degraded starch, e.g. maltodextrin, hydrophobic starch, amylose,starch-diethylaminoethylether and starch-2-hydroxyethylether, preferablyfrom β- or γ-cyclodextrin (Wacker Chemie; Gattfoseé).

[0047] In a preferred embodiment the bile acid sequestrant is acellulose or dextran derivative, e.g. DEAE-cellulose,guanidinoethylcellulose, and DEAE-Sephadex, preferably DEAE-cellulose.

[0048] In another preferred embodiment of the present invention, thestarch or starch derivative is selected from the group consisting of β-or γ-cyclodextrin, retrograded and/or degraded starch, hydrophobicstarch, amylose, starch-diethylaminoethylether andstarch-2-hydroxyethylether, preferably β- or γ-cyclodextrin.

[0049] In a further embodiment of the present invention, the bile acidsequestrant is selected from the group consisting of cholestyramine,colestipol, colesevelam, colestimide, sevelamer, cellulose,DEAE-cellulose, guanidinoethylcellulose, and DEAE-Sephadex, starch, β-or γ-cyclodextrin, preferably cholestyramine, colestipol, colesevelam,colestimide, sevelamer, DEAE-cellulose, and β- or γ-cyclodextrin, morepreferably cholestyramine, colestipol, sevelamer, DEAE-cellulose, and β-or γ-cyclodextrin, most preferably cholestyramine, colestipol, andsevelamer.

[0050] In a preferred embodiment of the present invention, the bile acidsequestrant is cholestyramine. In a further preferred embodiment, thebile acid sequestrant is colestipol. In a additional embodiment the bileacid sequestrant is sevelamer.

[0051] Pharmaceutical compositions incorporating both a compound of alipase inhibitor and a bile acid sequestrant are important embodimentsof the present invention. Such pharmaceutical compositions comprise atherapeutically effective amount of each of the compounds. Each dosageunit can obtain the daily doses of both compounds or may contain afraction of the daily dose, such as one-third of the doses.Alternatively, each dosage unit may contain the entire dose of one ofthe compounds, and a fraction of the dose of the other compound. In suchcase the patient would daily take one of the combination dosage units,and one or more units containing only the other compound.

[0052] Particularly, the above composition refer to compositionscomprising a) about 5 to about 1000 mg lipase inhibitor and b) about 0.1to about 20 g bile acid sequestrant. The compositions may comprise apharmaceutically acceptable excipient, e.g. a diluent or carrier. Thepharmaceutically acceptable excipient may be selected from the groupconsisting of fillers, e.g. sugars and/or sugar alcohols, e.g. lactose,sorbitol, mannitol, maltodextrin, etc.; surfactants, e.g. sodium laurylsulfate, Brij 96 or Tween 80; disintegrants, e.g. sodium starchglycolate, maize starch or derivatives thereof; binder, e.g. povidone,crosspovidone, polyvinylalcohols, hydroxypropylmethylcellulose;lubricants, e.g. stearic acid or its salts; flowability enhancers, e.g.silicium dioxide; sweeteners, e.g. aspartame; and/or colorants, e.g.β-carotene.

[0053] A preferred composition may comprise a) about 5 to about 1000 mglipase inhibitor; b) about 0.1 to about 20 g bile acid sequestrant; andoptionally pharmaceutically acceptable excipients selected from thegroup of about 0.1 to about 10 g fillers, about 0.05 to about 3.0 gsurfactant, about 0.05 to about 2.0 g disintegrant, about 0.02 to about2.0 g binder, about 0.001 to about 1.0 g lubricant, about 0.1 to about5.0 g flowability enhancer, about 0.01 to about 4.0 g sweetener, andabout 0.001 to about 0.5 g colorant. The preferred lipase inhibitor isorlistat.

[0054] In particular, the invention refers to pharmaceuticalcompositions comprising orlistat, a pharmaceutically acceptable bileacid sequestrant in conjunction with a pharmaceutically acceptableexcipient, e.g. a diluent or carrier, preferably to compositions whereinthe pharmaceutically acceptable bile acid sequestrant is selected fromas defined above. More preferably the compositions containing both alipase inhibitor, e.g. orlistat, and a bile acid sequestrant asdescribed above may comprise 5 to 1000 mg lipase inhibitor, preferablyabout 10 to about 500 mg lipase inhibitor, more preferably about 20 toabout 100 mg lipase inhibitor, e.g. orlistat. The preferred amounts fororlistat are about 10 to about 360 mg, preferably about 30 to about 120mg and most preferably about 40 to about 80 mg.

[0055] The pharmaceutical compositions may contain about 0.1 to about 20g bile acid sequestrant, preferably about 0.5 to about 10 g, and mostpreferably about 1 to about 5 g. The preferred bile salt sequestrant ischolestyramine.

[0056] The use according to the invention comprises the simultaneous,separate or sequential administration of lipase inhibitor and bile saltsequestrant.

[0057] The invention also refers to a process for preparing acomposition as described above, comprising mixing a lipase inhibitor ora pharmaceutically acceptable salt thereof with a bile acid sequestrantor a pharmaceutically acceptable salt thereof and one or morepharmaceutically acceptable excipient, e.g. a diluent and/or carrier.

[0058] Oral dosage forms are the preferred compositions for use in thepresent invention and these are the known pharmaceutical forms for suchadministration, for example tablets, capsules, bars, sachets, granules,syrups and aqueous or oily suspensions. The pharmaceutically acceptableexcipients (diluents and carriers) are known in the pharmacist's art.Tablets may be formed from a mixture of the active compounds withfillers, for example calcium phosphate; disintegrating agents, forexample maize starch, lubricating agents, for example magnesiumstearate; binders, for example microcrystalline cellulose orpolyvinylpyrrolidone and other optional ingredients known in the art topermit tabletting the mixture by known methods. Similarly, capsules, forexample hard or soft gelatin capsules, containing the active compoundwith or without added excipients, may be prepared by known methods. Thecontents of the capsule may be formulated using known methods so as togive sustained release of the active compound. For example, the tabletsand capsules may conveniently each contain the amounts of lipaseinhibitor and bile acid sequestrant as described above.

[0059] Other dosage forms for oral administration include, for example,aqueous suspensions containing the active compounds in an aqueous mediumin the presence of a non-toxic suspending agent such as sodiumcarboxymethylcellulose, and oily suspensions containing the activecompounds in a suitable vegetable oil, for example arachis oil. Theactive compounds may be formulated into granules with or withoutadditional excipients. The granules may be ingested directly by thepatient or they may be added to a suitable liquid carrier (e.g. water)before ingestion. The granules may contain disintegrants, e.g. aneffervescent pair formed from an acid and a carbonate or bicarbonatesalt to facilitate dispersion in the liquid medium.

[0060] In the compositions of the present invention the active compoundsmay, if desired, be associated with other compatible pharmacologicallyactive ingredients. Optionally vitamin supplements may be administeredwith the compounds of the present invention.

[0061] Both compounds, the lipase inhibitor and the bile acidsequestrant may be administered simultaneously, separately orsequentially. Preferably, the compounds or compositions are administeredduring a meal or 1-2 hours before or after a meal. The amount of bileacid sequestrant to be administered will depend on a number of factorsincluding the age of the patient, the severity of the condition and thepast medical history of the patient and lies within the discretion ofthe administering physician. For example, β- and γ-cyclodextrin (starchderivatives), cholestyramine, colestipol (amine containing polymer) anddiethylaminoethylcellulose (cellulose or dextran derivative) could beadministered 0.1-20 g per day, preferably 1-10 g per day, starch,amylose and other bile acid sequestrants described above 1-20 g per day.

[0062] The invention also provides the use of the above combination ofcompounds in the manufacture of a medicament for the prevention andtreatment of diseases associated with high plasma cholesterol levels,i.e. hypercholesterolemia. Additionally, it provides the combination andabove compositions for use in the prevention and treatment of diseasesassociated with high plasma cholesterol levels, i.e.hypercholesterolemia.

[0063] The invention also refers to a kit for prevention and treatmentof diseases associated with high plasma cholesterol levels, i.e.hypercholesterolemia, said kit comprising a first component which is alipase inhibitor and b) a second component which is a bile acidsequestrant in suitable oral unit dosage forms preferably comprising a)from 1 to 100 doses units of orlistat and b) from 1 to 100 doses unitsof a bile acid sequestrant.

[0064] Further, the present invention refers to the use of a lipaseinhibitor, e.g. orlistat, in the manufacture of a medicament for thetreatment of diseases associated with high plasma cholesterol levels,i.e. hypercholesterolemia in a patient who is also receiving treatmentwith a bile acid sequestrant or a pharmaceutically acceptable saltthereof.

[0065] A further embodiment of the present invention refers to a methodof treatment of diseases associated with high plasma cholesterol levels,i.e. hypercholesterolemia in a human in need of such treatment whichcomprises administration to the human of a therapeutically effectiveamount of a lipase inhibitor, e.g. orlistat, and a therapeuticallyeffective amount of a bile acid sequestrant or a pharmaceuticallyacceptable salt thereof. The administration of these compounds may beperformed simultaneously, separately or sequentially.

[0066] A further embodiment of the present invention refers to a lipaseinhibitor, e.g. orlistat, and a bile acid sequestrant andpharmaceutically acceptable salts thereof for simultaneous, separate orsequential use for the diseases associated with high plasma cholesterollevels, i.e. hypercholesterolemia. In addition the invention comprises alipase inhibitor, e.g. orlistat, and a bile acid sequestrant as definedabove and pharmaceutical salts thereof as a combined preparation forsimultaneous, separate or sequential use for prevention and treatment ofdiseases associated with high plasma cholesterol levels.

[0067] The invention will be better understood by reference to thefollowing examples which illustrate but do not limit the inventiondescribed herein.

EXAMPLES Example 1 Study

[0068] The effect of a combination of Orlistat/XENICAL® and a bile acidsequestrant cholestyramine/QUANTALAN® on plasma cholesterol wasexamined.

[0069] Three hyper-cholesterolemic volunteers, one female, 2 males,started on Orlistat/XENICAL® b.i.d. for 4 weeks to establish a stableplasma cholesterol level: They continued on a combination ofOrlistat/XENICAL® b.i.d. and cholestyramine/QUANTALAN® 4 g b.i.d. for afurther 4 weeks, which was followed by a 4 week washout period. Theeffect of this treatment schedule on plasma cholesterol is reported intable 1.

[0070] The plasma cholesterol of all three individuals is reasonablystable at the end of 6 weeks treatment with Xenical b.i.d. Afteraddition of Quantalan b.i.d. to the Xenical b.i.d. treatment plasmacholesterol clearly is reduced further by nearly 20%. The decrease isundoubtedly related to the Quantalan treatment: Upon cessation ofQuantalan ingestion plasma cholesterol levels return to the values onXenical alone.

[0071] This experiment demonstrates that the combined intake of a lipaseinhibitor and a bile acid sequestrant achieve a plasma cholesteroldecrease which is superior to the plasma cholesterol as produced by theintake of the lipase inhibitor alone an. i.e. the combined activity isat least additive.

[0072] Cholestyramine is known to enhance cholesterol synthesis byenhancing cholesterol degradation. In this sense Xenical as a (probable)cholesterol synthesis reducing compound and combined to bile acidsequestrants as cholesterol degradation enhancer represent asurprisingly active principle of plasma cholesterol reduction. TABLE 1Effect of Orlistat/XENICAL ® and Orlistat/XENICAL ® combined tocholestyramine/QUANTALAN ® on plasma cholesterol in 3 healthy volunteersplasma cholesterol (mg/100 ml) volunteer Xenical (Xenical + Quantalan)Xenical treatment b.i.d. b.i.d. b.i.d. week before 4 5 6 7 8 9 11 13 1232 252 239 233 — 198 ** 247 — 2 * 236 212 232 198 178 193 230 226 3 350269 — 277 246 241 262 282 300 mean 252 225 247 222 206 228 253 263 ±SD17 19 26 34 32 49 27 52

Example 2 Orlistat Pharmaceutical Compositions

[0073] A) Quantity Ingredient mg/Capsule orlistat 120.00microcrystalline cellulose (AVICEL PH-101)  93.60 sodium starchglycolate (PRIMOJEL)  7.20 sodium lauryl sulfate  7.20polyvinylpyrrolidone (Povidone (K-30))  12.00 purified Water* — talc 0.24 Total 240.24 mg

[0074] Procedure:

[0075] 1. Blend orlistat, microcrystalline cellulose, and sodium starchglycolate in a suitable mixer.

[0076] 2. Granulate with a solution of polyvinylpyrrolidone and sodiumlauryl sulfate in purified water.

[0077] 3. Pass the granulation through an extruder and pass theextrudate through a spheronizer to form pellets.

[0078] 4. Dry the pellets at 30° C.

[0079] 5. Add talc and mix.

[0080] 6. Fill into hard gelatin capsules.

[0081] B) Ingredient Quantity mg/Capsule orlistat  60 microcrystallinecellulose  46.8 sodium starch glycolate  3.6 sodium lauryl sulfate  3.6polyvinylpyrrolidone  6.0 purified water* — talc  0.12 Total 120.12 mg

[0082] Procedure:

[0083] 1. Blend orlistat, microcrystalline cellulose, and sodium starchglycolate in a suitable mixer.

[0084] 2. Granulate with solution of polyvinylpyrrolidone and sodiumlauryl sulfate in purified water.

[0085] 3. Pass the granulation through an extruder and pass theextrudate through a spheronizer to form pellets.

[0086] 4. Dry the pellets at 30° C.

[0087] 5. Add talc and mix.

[0088] 6. Fill into hard gelatin capsules.

[0089] C) Ingredient Quantity mg/Capsule orlistat  60 120 lactose  40 80 microcrystalline cellulose  60 120 sodium lauryl sulfate  5.7  11.4sodium starch glycolate  20  40 polyvinylpyrrolidone  10  20 purifiedwater* — — talc  0.2  0.4 Total 195.9 mg 391.8 mg

[0090] Procedure:

[0091] 1. Blend orlistat, lactose, microcrystalline cellulose and sodiumstarch glycolate in a suitable mixer.

[0092] 2. Granulate with a solution of polyvinylpyrollidone and sodiumlauryl sulfate in purified water.

[0093] 3. Pass the granulation through an extruder, and pass theextrudate through a spheronizer to form pellets.

[0094] 4. Dry the pellets at 30° C.

[0095] 5. Add talc and mix.

[0096] 6. Fill into hard gelatin capsules.

Example 3 Bile Acid Sequestrant Pharmaceutical Compositions

[0097] Quantity Ingredient mg/Capsule cholestyramine    4 g siliciumDioxide 0.495 g aspartame  0.05 g β-carotene 0.001 g purified water* —Total  4.5 g

[0098] Procedure:

[0099] 1. Blend colestyramine, and silicium dioxide in a suitable mixer.

[0100] 2. Granulate with a solution/colloidal suspension of aspartameand beta-carotene in purified water.

[0101] 3. Pass the granulation through an sieve.

[0102] 4. Dry the granules at 60° C.

[0103] 5. Pass the dry granulation through an sieve

[0104] 6. Fill into sachets.

Example 4 Bile Acid Sequestrant Pharmaceutical Compositions

[0105] Quantity Ingredient mg/Capsule cholestyramine    4 g siliciumdioxide  0.5 g saccharose    3 g β-carotene 0.001 g purified water* —total  7.5 g

[0106] Procedure:

[0107] 1. Blend colestyramine, silicium dioxide, and saccharose in asuitable mixer.

[0108] 2. Granulate with a solution/colloidal suspension of aspartameand beta-carotene in purified water.

[0109] 3. Pass the granulation through an sieve.

[0110] 4. Dry the granules at 60° C.

[0111] 5. Pass the dry granulation through an sieve

[0112] 6. Fill into sachets.

Example 5 Bile Acid Sequestrant Pharmaceutical Compositions

[0113] Quantity Ingredient mg/Capsule cholestyramine    4 g aspartame 0.5 g β-carotene 0.001 g purified water* — Total  4.05 g

[0114] Procedure:

[0115] 1. Fill colestyramine in a suitable mixer.

[0116] 2. Granulate with a solution/colloidal suspension of aspartameand beta-carotene in purified water.

[0117] 3. Pass the granulation through an sieve.

[0118] 4. Dry the granules at 60° C.

[0119] 5. Pass the dry granulation through an sieve

[0120] 6. Fill into sachets.

Example 6 Orlistat/Bile Acid Sequestrant Pharmaceutical Compositions

[0121] Quantity Ingredient mg/Capsule orlistat   120 mg maltodextrinum  740 mg cholestyramine  4000 mg aspartame   440 mg purified water* —total  5.3 g 

[0122] Procedure:

[0123] 1. Melt orlistat in a mixer and add maltodextrin.

[0124] 2. Mix until solidification at room temperature (first part)

[0125] 3. Add cholestyramine and mix

[0126] 4. Granulate with a solution/colloidal suspension of aspartame inpurified water.

[0127] 5. Pass the granulation through an sieve.

[0128] 6. Dry the granules at 60° C.

[0129] 7. Pass the dry granulation through an sieve (second part)

[0130] 8. Blend both parts in a mixer

[0131] 9. Fill into sachets.

Example 7 Orlistat/Bile Acid Sequestrant Pharmaceutical Compositions

[0132] Quantity Ingredient mg/Capsule orlistat   120 mg microcrystallinecellulose   240 mg sodium starch glycolate   60 mg sodium lauryl sulfate  30 mg crospovidone   50 mg cholestyramine  4000 mg aspartame   200 mgpurified water* — total  5.2 g 

[0133] Procedure:

[0134] 1. Blend cholestyramine, orlistat, microcristalline cellulose(Avicel), sodium starch glycolate and crospovidone in a suitable mixer.

[0135] 2. Granulate with a solution/colloidal suspension of sodiumlauryl sulfate, aspartame in purified water.

[0136] 3. Pass the granulation through an sieve.

[0137] 4. Dry the granules at 30° C.

[0138] 5. Pass the dry granulation through a sieve

[0139] 6. Fill into sachets.

Example 8 Orlistat/Bile Acid Sequestrant Pharmaceutical Compositions

[0140] Quantity Ingredient mg/Capsule orlistat    4 g orlistat   120 mgmaltodextrinum   740 mg colestipol  4000 mg aspartame   440 mg purifiedwater* — total  5.3 g 

[0141] Procedure:

[0142] 1. Melt orlistat in a mixer and add maltodextrin.

[0143] 2. Mix until solidification at room temperature (first part)

[0144] 3. Add colestipol and mix.

[0145] 4. Granulate with a solution/colloidal suspension of aspartame inpurified water.

[0146] 5. Pass the granulation through an sieve.

[0147] 6. Dry the granules at 60° C.

[0148] 7. Pass the dry granulation through an sieve (second part)

[0149] 8. Blend both parts in a mixer

[0150] 9. Fill into sachets.

Example 9 Orlistat/Bile Acid Sequestrant Pharmaceutical Compositions

[0151] Quantity Ingredient mg/Capsule orlistat   120 mg microcrystallinecellulose   240 mg sodium starch glycolate   60 mg sodium lauryl sulfate  30 mg crospovidone   50 mg colestipol  4000 mg aspartame   200 mgpurified water* — total  5.2 g 

[0152] Procedure:

[0153] 7. Blend colestipol, orlistat, microcrystalline cellulose(Avicel), sodium starch glycolate and crospovidone in a suitable mixer.

[0154] 8. Granulate with a solution/colloidal suspension of sodiumlauryl sulfate, aspartame in purified water.

[0155] 9. Pass the granulation through an sieve.

[0156] 10. Dry the granules at 30° C.

[0157] 11. Pass the dry granulation through a sieve

[0158] 12. Fill into sachets.

[0159] Upon reading this specification, various alternative embodimentswill become obvious to the skilled artisan. These variations are to beconsidered within the scope and spirit of the subject invention, whichis only to be limited by the claims that follow and their equivalents.

What is claimed is:
 1. A method of reducing plasma cholesterol levels ina hypercholesterolemic patient, which comprises administering to thepatient a therapeutically effective amount of a lipase inhibitor and atherapeutically effective amount of a bile acid sequestrant selectedfrom the group consisting of cholestyramine, colestipol, colesevelam,colestimide, sevelamer, DEAE-cellulose, guanidinoethylcellulose,DEAE-Sephadex, β-cyclodextrin, and γ-cyclodextrin.
 2. The methodaccording to claim 1, wherein the lipase inhibitor is orlistat.
 3. Themethod according to claim 1, wherein the bile acid sequestrant isselected from the group consisting of DEAE-cellulose,guanidinoethylcellulose, and DEAE-Sephadex.
 4. The method according toclaim 1, wherein the bile acid sequestrant is selected from the groupconsisting of cholestyramine, colestipol, sevelamer, DEAE-cellulose,β-cyclodextrin, and γ-cyclodextrin.
 5. The method according to claim 4,wherein the bile acid sequestrant is selected from β-cyclodextrin orγ-cyclodextrin.
 6. The method according to claim 1, wherein thetherapeutically effective amount of a lipase inhibitor and atherapeutically effective amount of a bile acid sequestrant areadministered as a medicament that comprises about 5 mg to about 1000 mglipase inhibitor, about 0.1 g to about 20 g bile acid sequestrant, andone or more pharmaceutically acceptable excipients.
 7. The methodaccording to claim 6, wherein the pharmaceutically acceptable excipientis selected from the group consisting of fillers, sugars, sugaralcohols, a mixture of one or more sugars and sugar alcohols,surfactants, disintegrants, binders, lubricants, flowability enhancers,sweeteners, and colorants.
 8. The method according to claim 7, whereinthe medicament comprises: a) about 5 mg to about 1000 mg lipaseinhibitor; b) about 0.1 g to about 20 g bile acid sequestrant; and apharmaceutically acceptable excipients selected from the groupconsisting of about 0.1 to about 10 g fillers, sugars, sugar alcoholsand mixtures thereof, about 0.05 g to about 3.0 g surfactants, about0.05 g to about 2.0 g disintegrants, about 0.02 g to about 2.0 g binder,about 0.001 g to about 1.0 g lubricants, about 0.1 g to about 5.0 gflowability enhancers, about 0.01 g to about 4.0 g sweeteners, and about0.001 g to about 0.5 g colorants.
 9. The method according to claim 6,wherein the medicament comprises about 10 mg to about 500 mg lipaseinhibitor.
 10. The method according to claim 9, wherein the medicamentcomprises about 20 mg to about 100 mg lipase inhibitor.
 11. The methodaccording to claim 9, wherein the medicament comprises about 10 mg toabout 360 mg orlistat.
 12. The method according to claim 11, wherein themedicament comprises about 30 mg to about 120 mg orlistat.
 13. Themethod according to claim 12, wherein the medicament comprises about 40mg to about 80 mg orlistat.
 14. The method according to claim 6, whereinthe medicament comprises about 0.5 g to about 10 g bile acidsequestrant.
 15. The method according to claim 14, wherein themedicament comprises about 1 g to about 5 g bile acid sequestrant. 16.The method according to claim 1, wherein the lipase inhibitor and bileacid sequestrant are administered simultaneously.
 17. The methodaccording to claim 1, wherein the lipase inhibitor and bile acidsequestrant are administered separately.
 18. The method according toclaim 1, wherein the lipase inhibitor and bile acid sequestrant areadministered sequentially.
 19. The method according to claim 1, whereinthe lipase inhibitor is selected from the group consisting of orlistat,2-decyloxy-6-methyl-4H-3,1-benzoxazin-4-one,6-methyl-2-tetradecyloxy-4H-3,1-benzoxazin-4-one, and2-hexadecyloxy-6-methyl-4H-3,1-benzoxazin-4-one.